A Review of Isothermal Amplification Methods and Food-Origin Inhibitors against Detecting Food-Borne Pathogens

Abstract

The isothermal amplification method, a molecular-based diagnostic technology, such as loop-mediated isothermal amplification (LAMP) and recombinase polymerase amplification (RPA), is widely used as an alternative to the time-consuming and labor-intensive culture-based detection method. However, food matrices or other compounds can inhibit molecular-based diagnostic technologies, causing reduced detection efficiencies, and false-negative results. These inhibitors originating from food are polysaccharides and polyphenolic compounds in berries, seafood, and vegetables. Additionally, magnesium ions needed for amplification reactions can also inhibit molecular-based diagnostics. The successful removal of inhibitors originating from food and molecular amplification reaction is therefore proposed to enhance the efficiency of molecular-based diagnostics and allow accurate detection of food-borne pathogens. Among molecular-based diagnostics, PCR inhibitors have been reported. Nevertheless, reports on the mechanism and removal of isothermal amplification method inhibitors are insufficient. Therefore, this review describes inhibitors originating from food and some compounds inhibiting the detection of food-borne pathogens during isothermal amplification.

Performance of Loop-Mediated Isothermal Amplification Technique in Milk Samples for the Diagnosis of Bovine Tuberculosis in Dairy Cattle Using a Bayesian Approach

“…Moreover, Zhang et al demonstrated that the LAMP test can detect the DNA of M. bovis without any cross-reaction with other mycobacterial DNA, indicating a high Sp of the test [ 22 ]. However, the performance of the LAMP test may be affected by the high concentration of calcium ions in milk, which can competitively bind to DNA polymerases with magnesium ions, a necessary cofactor for the reaction [ 23 ]. Additionally, prior estimates of disease prevalence given in the current study are based on the reports of previous surveys using only the SIT test.…”

Development of a PMA‐LAMP visual detection assay for viable Cronobacter sakazakii

“…The complex components of food samples (Moon et al . 2022) as well as other pathogens in food might inhibit the nucleic acid amplification. Therefore, the artificially contaminated samples were detected by the PMA‐LAMP‐based visual assay developed in this study to determine the reliability.…”

Development of Loop-Mediated Isothermal Amplification (LAMP) Assays for the Rapid Authentication of Three Swimming Crab Species

“…The more the primers in the LAMP reaction, the higher the quality and integrity of the template required [ 29 , 30 ]. Moreover, seafood has been reported to contain some types of Bst polymerase inhibitors such as cationic polysaccharides and/or calcium ions that inhibit polymerase activity and competitively bind to Bst polymerases with magnesium ions [ 31 ]. Therefore, a DNA extraction method developed for food samples is required to obtain high-quality DNA [ 32 ].…”